Concentration of fat-soluble vitamins



; Patented Dec. 17, 1946 UNITED STATE s 1 PATENT! OFFICE CONCENTRATIONor FAT-SOLUBLE VITAMINS Loran Old Buxton, East Orange, N. 1., asslgnorto I National Oil Products Company, Harrison, N. 1., a corporation ofNew Jersey No Drawing. Application August 4, 1942,

Serial No.453,560

Claims. l This invention relates in generalto the pro 'duction offat-soluble vitamin concentrates and in particular to a process ofproducing highly- 1 potent concentrates of vitamin A.

. Fat-soluble vitamin concentrates are being-proa duced at the presenttime by many different processes, the basic principle of the most commoncommercial procedures involving the complete saponiilcation of thesaponiflable matter in a fat-soluble vitamin containing oil and therecovery of the unsaponiflablematter from the resuiting soaps. Manyattempts have been made and proposals offered to improve the foregoingsaponification processes with a view toward increasing the potency ofthe ultimate concentrate.

One such process involves the extraction of the original oil withethanol at room temperature and the subsequent saponiflcation of theethanol extracted fraction. The latter process and many others whichhave been proposed, including high vacuum distillation, have fallenshort in providing highly potent concentrates of vitamin A.

It is the object of this invention to provide an improved process forproducing concentrates of fat-soluble vitamins.

Another object of the invention is to provide an improved process forproducing concentrates of vitamin A.

Another object of the invention is to provide highly potent concentratesof vitamin A.

Other objects of the invention will in part be obvious and will in partappear hereinafter.

I have discovered that the above and other objects of the invention maybe realized by extracting a fat-soluble vitamin-containing marine oilwith a solvent which is characterized by being miscible with the oil attemperatures above room temperature and partially immiscib e therewithat temperatures below room tem erature. saponifying thesolvent-extracted fraction and recovering the unsaponifled matter fromthe resulting soaps. Preferably-several extractions of the fat-solublevitamin-containing marine oil are,

.2 after disclosed, and the scopeof the invention will be indicated inthe claims.

The first basic-step of the process invoives tlie j extraction of theoriginal oil with a particular class of solvents hereinafter described.The sol vent employed in accordance with the process of the inventionmay be selected from a large number of aliphatic solventsfound to beuseful as a result of extensive experimentatiom' the choice of thesolvent will depend to some extent upon the properties of the oil{ to betreated. as will become more evident from'therletailed descrip- "tionhereinafter given.

that the solventspreferably employed are mem- Results have indicatedbers of well recognized chemical classes; it has. also been found thatthe-number of carbon atoms in the solvent to be used is. a particularlyimportant' factor in determining the availability thereof for use in thepractice ,of this invention. i I

The following table sets forth the classes of-sol- V vents which havebeen found to be particularly 7 4. Aliphatic ketones containing not moruseful in the practice of this invention:

Table I l. Aliphatic and alicyclic monohydroxy alcohols Q containingfrom 3 to 6 carbon atoms.- 2. Esters formed by the reaction of aliphaticand alicyciic alcohols with aliphatic monocarboxyllc acids, said esterscontaining not more I than. 8 carbon atoms.

3. Aliphatic and aiicyclic aldehydes containing not more than 6 carbonatoms.

than 8 carbon. atoms.

Solvents falling in the classes above listed are all liquid aliphaticorganic compounds characterized by being miscible with fat-solublevitamincontalnlng marine oils at temperatures above room temperature, 1.e.20 to 25 C., and partially immiscible therewith at temperatures belowroom temperature, and it has been found that solvents falling withinthese classes of compounds list thereof is given herewith; it is to beunder stood, however, that thislistis not intended to may be used in thepractice of this invention.

In addition it will be noted that the preferred solvents possessrelatively low freezing points.-

In order to more'full'y illustrate the nature of the solvents which maybeemployed, a partial be complete, but is merely illustrative of thesolvents which may be employed. Thus it has been found that thefollowing solvents may be used: n-propyl alcohol, isopropyl alcohol,isopropylamine. n-butyl' alcohol, n-amyl alcohol, isoamyl 7 alcohol.secondary amyl alcohoLfurfuryl alcohol.

- 3 allyl alcohol. diacetone alcohol, e-hydroxy ethyl acetate, me hylformate, ethyl formate, ethyl acetate. methyl acetate, isopropylacetate. glycol diformate. glycol diacetate, methyl levulinate,

ethyl levulinate. methyl aceto acetate, ethyl aceto acetate. methylfuroate} vinyl acetate, furfural pron onaldehyde, crotonaldehyde,acetone, methyl ethyl ketone, acetonyl acetone and propylenechlorhydrin. Mixtures of .these solvents may also be used. It will benoted that all thesesolvents belong to that class of aliphatic organiccompounds which has the property of, being miscible with fat-soluble vitmin-conta ning marine oils at temperatures above room temperature andpartially immiscible therewith at tempera tures below room temperature;

have relatively low freezing points.

Occasionally it may be found that certain of I the solvents'hereinabovementioned may be-toofurthermore, it i will be noted that the majority ofthese solvents miscible with some of the oils]v which may be treated bythis invention :to: effect .a separat on;

of highly potent: vita-min fractions therefrom;

thus, for examnlaacetone is toomiscible with some fat-solublevitamin-containing oil to ac ever. this condition can beeasilyacorrectedby diluting the solvent either with a smallarnount ofwaterorwith'a liquid aliphatic' organicsolvent relatively immisciblewith fatty oils. In; gen- 7 compllsh the purposes'ofzthisinvention.,-HoW.-.

vitamin A at temperatures above this value. It is preferred to form thesolution of oil in the solvent by first heating the solvent to be usedto a predetermined temperature at which the oil to be added'willsubstantially completely dissolve in the solvent, and then adding theoil to the solvent with agitation-the operatio'n being carriedatmosphere.

out inan inert gas the vitamin-containing oil in The solution of the isolvent prepared as hereinabove described may then, in accordance withthe process of the invention, be permitted to cool so as to efi'ect aseparation of the solution of the highly potent vitaminextr'actfrom' theremainder 'of the oil. The temperature to which the solution is cooledmay'vary from about room temperature to as 1 low as--'70 C. or lower. Ithas been found, howw at r it ,iSprefietable to cool the solution withagitation to temperatures somewhat below about 0C., e. g. in theneighborhood of 18 C. Upon .cooling, the solution separates into twolayers,

one layer composed chiefly of the portion of .the

original oil insoluble in the solvent atlow tem-- T peratures and .1

solvent solution, of a highly potentvitamin fraction. The solvent layerobtained upon coolingthe the other layer composed of the solution may befiltered and'then treated to reeral it may be said that the effect ofdiluting-any of the above solvents with water will be to-render thesolvents more immiscible-with fatty oils, so

that if difficulty is encountered in effecting-Proper Vvitamin-containing oils, this. difllcultycangenseparation ofrthe highlypotent extracts fromthe erally be overcome 'by the addition of a smallamount of waterte the solvent.

The solvents preferably employed. intheprac- 1' tice of theinventionarethe aliphatic' alcoholsr In most containing from 3'to 6. carbon atoms:of these I solvents isopropanol and diacetone alcohol ha e proved to bethe most successful. The presence of the hydroxyl group seems to impartto these:

solvents properties which make them particularly useful for the presentpurposes; whether this fac- "movethe' soIvent-thereIrOm-Le. g. by vacuum(115- tillation, whereby-an oil is recovered having a vitamin potencyfar inexce'ss'of that of the original oil;,thepe'rcent'age increase invitamin po- "tency may be about 400%.

anywhere between about 50 and 3 to 6 more" extractions The firstextractwhich is obtained is preferably 1 separately from tor is duej to'some activating influence possessed, by this group is not known, but itis believed that the presence ofthe hydroxyl group in such.solventsmakes them more capable' of'extracting vitaminA esters from oilscontainin'g such esters.

In carrying out the process of the invention thefat-solublevitamin-containing marine oil tobe treatedis first mixed with anyparticular solvent of the aforementioned type. The oil treated may i beany of the marine oils containing. vitaminA with orwithout vitamin D,such as, for'exa'mpla' cod liver :oil, shark liver oil, tuna liver oil,hall? but liver oil,;mackerelliver oil,.ling cod liver'oil, soleliveroil, spear; fishliver oil, sword fish liver oil, sardine. oil,herring oil, menhadenoil, whale 'liver oil, etc. The relativeproportion'of oil to solvent in the mixture may varywidely';preferably'the ratio of. solvent tooil should be greater than-one andinmost cases mixtures containing,"

a the fractions from the between about 2% and,about 25% oil are'm'ost isuitable. Thismixture maythenbe heated until I the oil orthegreater partthereof is dissolved in the solvent. The temperature. to which themixture of oil an solvent is heated-may vary widely depending upon thenature of the in redients 7 contained in .the mixture; in general itmaybe stated that it is inadvisable to heat fat-solublevitamin-containingoils to temperatures-inexcess of 175 c. because of therelative instability of possible by the "vitamin A concentrates on'hcommercial scale isram, i; e. essentially pure sa'pom'iled and thevitamins recoveredtherefrom the other extracts. The; second extracts mayeither-be combined and subsequent and then saponified and the vitaminconcentrate recovered therefrom or each extract may be saponifiedseparately and the unsaponiflable matter then recovered from eachsaponifled extract. I have found'th'at invariably. the firstextractcontains the lowest ratio of vitamins inrelation to thtotalunsaponifiable matter when compared to the 'subsequentextracts ofthe same 1 oil.

In many cases given oil. In view thereof, it is preferable, particularlywhen exceedingly high potent vitamin A concentrates are desired, tosaponify each frac- I tion separately. In most cases, however, all of vseconclon may be-combined priorto saponification as the first extractappears to'contain a much higher ratio of un-. saponiflable matter tovitamlnA than each of the succeeding fractions. in any event, however.the firstrfractio'n-will yield a vitamin A concentrate y y having ahigher potency than a concentrate prepared by directly saponifying'thesame 011.; It is processof the invention to obtain having a potency ofabove 3,000,000 units per --18 0;; this method a continuous extractionof a highly potentjvitaminfraction'froin the oil. cases at least 2extractions of the fat-,.

soluble vitamin-containingoil should be made, are preferably made. 1

the ratio of vitamin). tothe 1 unsaponiflable matter increasessuccessively from the first to the sixthfraction extracted from avitamin-A; Inprder to determine whether or not to saponify the first andsubesequent extracts separately or together,

it is usually advisable to run laboratory tests on small samples of eachextract to determine the ultimate potencies to be expected. a

The saponification of the extracts and the recovery of theunsaponifiable material therefrom may be carried out in any suitablemanner. In f all cases, it is preferred, of course, that the ex;

tracts be recovered from the polar solvent solutions thereof before theyare saponified. Preferably the saponification of the extracts is carriedout by the process of application, Serial- No. 333,114 of Buxton andSimons filed May 3, 1940, which has issued as Patent No. 2,318,748, orthe process of application, Serial No. 350,166 of Buxton and Colman,filed August 2, 1940, which has issued as Patent No. 2,318,749, sincemuch more efficient recovery of the fat-soluble vitamins'will I beobtained.

For a fuller understanding of the nature and objects of theinvention,'reference should be had to the following examples which aregiven merely to further'illustrate the invention and are not to beconstrued in a limiting sense all parts given being by weight and allvitamin potencies being expressed inU. S. P. units.

Example I 100 parts of crude shark liver 011 containing 99,000 U. S. P.units of vitamin A per gram were mixed with 400 parts of 99% isopropanoland the mixture warmed to about C. to dissolve the oil. The solution wasthen cooled gradually to about 25 C. After about 48 hours the upperisopropanol layer was decanted and filtered. The

insoluble solidified oil fraction was re-extracted three more times asabove.- The four isopropanol extracts were combined, filtered and theisopropanol removed by distillation under reduced pressure and in thepresence of N2 gas.

100 parts of the combined isopropanol extracts containing 262,000 U. S.P. units of vitamin A per gram were mixed with parts of ethylenedichloride and 3 parts of 99% isopropanol. While stirring in thepresence 0f.N2 gas sufiicient 45% aqueous KOH (43 grams) to completelysaponify the oil was added. After about 20 minutes the stirring wasceased and the resulting thick supersolvented emulsion insulated andallowed to stand at room temperature over-night. The soap-mass was thenheated to about C. while stirring in the presence of N2 gas for about 30minutes to break the emulsion and to fiocculatethe soap.

400 parts of ethylene dichloride and sufficient water to bring themoisture content .of the soap up to about 24% were added and thesoap-mass cooled to room temperature. The stirring was stopped and afterabout 30 minutes the lower solvent layer removed. The flocculated soapparticles were then extracted six more times with 400 part portions ofethylene dichloride. The seven ethylene dichloride extracts werecombined, filtered and the ethylene dichloride removed by distillation.The recovered vitamin A alcohol concentrate contained 2,400,000 U; S. P.units of vitamin A per gram. A concentrate produced directly from thecrudeoil by the above saponiflcation process yielded a. productcontaining 1,600,- 000 U. S. P. units of vitamin A per gram.

' 100 parts of crude shark liver oil containing 106,000 U. s. P. unitsof vitamin A per gram were mixed with 4000 parts of 99% isopropanol andthe mixture warmed to about '33 C. to dissolve the oil. The solution wasthen cooled gradually to about 25 C. and after about 24 hours the upperisopropanol layer was decanted and filtered. The

insoluble solidified oil layer was re-extracted three more times asabove. Each isopropanol' extract fraction was freed of isopropanolseparately. by subjecting the same to distillation under reducedpressure in the" presence of N2 gas. The

following unsaponifiable and vitamin A"values 3 on the crude oil and onthe four separate isopropanol extract fractionsv bring out clearly whyunsaponifiable fractions prepared frorn'theisec ond, third and fourthextract fractions-are'materially more potent in vitamin A than similarlyprepared fractions from the crude oil'or fro the first isopropanolextract fraction.

Unsaponifled value mm A per Crude oil 5. 4 First isopropanol extract.13. 0 Second isopropanol extract.. 11.0 Third isopropanol extract. 8.8Fourth isopropanol extract 8. 4

20 parts of, each of the above samples were 7 completely saponified andthe unsaponifiable fraction recovered'as described'in Examplel'.

The following vitamin Aresults wereobt'ainedon the unsaponiflable'fractions. fromeach s pfj the aforesaid samples. 1 A g Units vitamin Aper gram on the unsaponifiable Crude i 9 0 First isopropanol extractSecond isopropanolextract 2,03%900- Third isopropanol extract 2 ,4 70,000 Fourth isopropanol extract 2,050,000

It is evident from the above description are-- examples that the processof the invention pro-q ployed to supply the vitamin needs of livestockand poultry by admixing the oil" with livestock and poultry feeds in,the usual manner. In case the vitamin potency of this oil is slightlybelow that desired for use in farm feeds, it may easily be adjusted tothe desired potency with concentrates of the necessary vitamin.

Since certain changes may be made in carrying I out the process withoutdeparting from the' scope of the invention, it is intended that allmatter contained in the above description shall be interpreted asillustrative and norm alimiting sense.

Having described my invention, what 'Iclainii as new and desireto'secureby-LettersPatent is;

'1. A process of producing an improved fath soluble vitamin concentrate,which comprises contacting a fat-soluble vitamin-containing marine oilwith a solvent selected from the group consisting of aliphatic andalicyclic monohydroxy alcohols containing from 3 to 6 carbon atoms,,

esters formed by the reacton of aliphatic and all- Highly potentconcentrates of fat cyclio alcohols with aliphatic monocarboxylic acids,said esters containing not more than 8 carbon atoms, aliphatic andalicyclic aldehydes containing not more than 6 carbon atoms andaliphatic ketones containing not more than 6 carbon atoms, heating themass to dissolve at least a major portion of the oil in the solvent,cooling the mass to a temperature within the range of C. to -70 C.whereby layers are donned. separating the solvent-soluble fraction fromthe mass, saponifying said soluble fraction and recovering theunsaponifled matter therefrom.

' whereby layers are formed, separating the solvent-soluble fractionfrom the mass, saponifying said soluble fraction and recovering theunsaponifled matter therefrom.

3. A process of producing an improved fatsoiuble vitamin concentrate,which comprises contacting a fat-soluble vitamin-containing marine oilwith isopropanol, heating the mixture to dissolve at least a majorportion of the oil in the isopropanol, cooling the mass to a temperaturewithin the range of 0 C. to 'l0 0. whereby layers are formed, separatingthe isopropanol-s'oiublo fraction from the mass, saponifying saidsoluble fraction'and recovering the unsaponiiled matter therefrom.

4. A process of producing an improved fat-solubie vitamin concentrate,which comprises contacting a fat-soluble vitamin-containing marine oilwith diacetone alcohol, heating the mixture to dissolve at least a majorportion of the oil in the diacetone alcohol, cooling the mass to atemperature within the range of 0 C. to -*70 C.

whereby layers are formed, separating the diacetone alcohol-solublefraction from the mass, saponifying said soluble fraction and recoveringthe unsaponified matter therefrom.

, 5. A process of producing an improved fat-soluble vitamin concentrate,which comprises contacting a iat-soluble vitamin-containing marine oilwith aqueous acetone, heating the mixture to dissolve at least a majorportion of the oil in the ,atoms, heating the mass to dissolve at leasta major portion of the oil in the solvent, cooling the mass to atemperature within the range of 0 C. to --70 C. whereby layers areformed, sep- 7 arating the solvent-soluble fraction from the mass,re-extracting at least twice more the solvent-insoluble fraction asabove, saponifying said soluble'fractio'ns and recovering theunsaponii'led matter therefrom.

7. A process of producing an improved fatsoluble vitamin concentrate,which comprises contacting a fat-soluble vitamin-containing marine oilwith a solvent selected from the group consisting of aliphatic andalicyclic monohydroxy aqueous acetone, cooling the mass to atemperacids, said esters containing not more than 8 caralcoholscontaining from 3 to 6 carbon atoms,

heating the mass to dissolve at least a major portion of the oil in thesolution, cooling the mass 4 therefrom.

8. A process of producing an improved fatsoluble vitamin concentrate,which comprises contacting a fat-soluble vitamin-containing marine oilwith isopropanol, heating the mixture to dissolve at least a majorportion of the oil in the isopropanol, cooling the mass to a temperaturewithin the range of 0 C. to C. whereby layers are formed, separating theisopropanolsoluble fraction from the mass, re-extracting at least twicemore the isopropanol-insoluble fraction as above, saponifying saidsoluble fractions and recovering the unsaponiiied matter therefrom.

9. A process of producing an improved fatsoluble vitamin concentrate,which comprises contacting a fat-soluble vitamin-containing marine oilwith diacetone alcohol, heating the mixture to dissolve at least a majorportion of the oil in the diacetone alcohol, cooling the mass to atemperature within the range of 0 C. to 70 C. whereby layers are formed,separating the diacetone alcohol-soluble fraction from the mass,re-extracting at least twice more the diacetone alcohol-insolublefraction as above, saponifying said soluble fractions and recovering theunsaponiiied matter therefrom.

10. A process of producing an improved fatsoluble vitamin concentrate,which comprises contacting a fat-soluble vitamin-containing marine oilwith aqueous acetone, heating the mixture to dissolve at least a majorportion of the oil in the aqueous acetone, cooling the mass to atemperature within the range of 0 C. to 70 C. whereby layers are formed,separating the aqueous acetone-soluble fraction from the mass,reextracting at least twice more the aqueous acetone-insoluble fractionas above, saponifying said soluble fractions and recovering theunsaponifled matter therefrom.

LORAN OID BUXTON.

